OP0195 TARGETED ABLATION OF KNEE-INNERVATING NOCICEPTORS HAS PROFOUND EFFECTS ON JOINT DAMAGE IN EXPERIMENTAL OSTEOARTHRITIS
Authors: L. Wang, S. Ishihara, J. LI, A. Obeidat, R. Miller, A. M. Malfait
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> Sensory afferents abundantly innervate the knee joint. The vast majority of these are pain-sensing nociceptors, which express the voltage-gated sodium channel, Na<sub>V</sub>1.8. Using Na<sub>V</sub>1.8Cre-tdTomato reporter mice, we have documented innervation changes that occur with experimental murine osteoarthritis (OA), including sprouting of nociceptors in the medial synovium and in subchondral bone [1]. Upon intra-articular (IA) injection into the mouse knee, adeno-associated virus serotype AAV.PHP.S is retrogradely transported to the lumbar dorsal root ganglia (DRG), where the cell bodies of sensory neurons reside [2]. <h3>Objectives</h3> We leveraged this technique to selectively ablate knee-innervating nociceptors by IA injection of AAV.PHP.S carrying diphtheria toxin A (dtA), and determine the effect on joint damage after destabilization of the medial meniscus (DMM). <h3>Methods</h3> Expt. 1: AAV.PHP.S containing pAAV-EF1a-mCherry-flex-dtA (AAV-dtA) was injected into the right knee of 7-week old male C57BL/6 Na<sub>V</sub>1.8Cre mice (3 µL, 10<sup>12</sup> vp/mL). Three weeks later, the TLR2 ligand, Pam3CSK4 (3 µg), was injected into these same knees, followed by knee hyperalgesia measurement (using PAM) over 24-hours [3]. Na<sub>V</sub>1.8Cre mice injected with AAV.PHP.S-EF1a-flex-GFP (AAV-Control) as well as wild type mice injected with AAV-dtA were included as controls (n=6 mice/group). Expt. 2: AAV-dtA or AAV-Control was injected IA into the right knees of 7-week old male Na<sub>V</sub>1.8Cre mice and 3 weeks later, DMM surgery was performed (n=19 mice/group). Mice were sacrificed 10 or 17 weeks after DMM, and knees collected for histologu using a modified OARSI scoring system (toluidine blue, frontal plane). Expt. 3: Ten-week old male Na<sub>V</sub>1.8Cre mice underwent DMM surgery, and 4 weeks later, AAV-dtA or AAV-Control was injected IA into the operated knees (n=12 mice/group). Knees were collected 17 weeks after surgery for histology. <h3>Results</h3> Expt. 1: Na<sub>V</sub>1.8Cre mice injected with AAV-Control and WT mice injected with AAV-dtA responded to TLR2 stimulation, showing markedly decreased knee pain thresholds between 1 and 6 hours after IA injection of Pam3CSK4, as described [3]. In contrast, Na<sub>V</sub>1.8Cre mice injected with AAV-dtA did not respond to Pam3CSK4, indicating functional effects of ablating knee-innervating nociceptors with AAV-dtA. Expt. 2: Ten weeks after DMM, all mice showed mild OA joint damage, with no difference between groups (Figure 1A). However, while AAV-Control injected joints showed markedly increased cartilage damage by week 17, AAV-dtA injected joints showed no progression of cartilage degeneration compared to week 10. Osteophytes were comparable between the groups. Expt. 3: Seventeen weeks after DMM, mice that had been injected with AAV-dtA 4 weeks <i>post</i> DMM showed markedly increased cartilage degeneration, total joint scores, and osteophyte sizes compared to AAV-controls (Figure 1B). <h3>Conclusion</h3> IA delivery of AAV-dtA into enables selective ablation of Na<sub>V</sub>1.8+ neurons in the knee, as is evident from ablated pain responses to TLR2 stimulation. Experiments in the DMM model revealed a striking effect of ablation of joint nociceptors on joint damage: Nociceptor ablation <i>before</i> DMM significantly <i>attenuated</i> cartilage degeneration in late stage OA, while ablation of knee nociceptors <i>after</i> DMM markedly <i>worsened</i> joint damage and osteophyte formation. These findings suggest an important role for nociceptors in joint homeostasis, with differential effects if nociceptors are ablated before or after joint damage occurs. Our findings suggest a neurogenic contribution to joint homeostasis. Current work focuses on documenting the joint nociceptive innervation in the different experimental groups, and the relationship with pain. <h3>References</h3> [1]Obeidat AM, <i>et al</i>. Osteoarthr Cart. 2019;27(11):1669-79. [2]Chakrabarti S, <i>et al</i>. Arthritis Rheum. 2020;72(10):1749-58. [3]Miller RE, <i>et al</i>. JCI Insight. 2018;3(6). <h3>Acknowledgements</h3> NIH/NIAMS grants R61AR073576, R01AR060364, R01AR064251, and P30AR079206. <h3>Disclosure of Interests</h3> Lai Wang: None declared, Shingo Ishihara: None declared, Jun Li: None declared, Alia Obeidat: None declared, Rachel Miller: None declared, Anne-Marie Malfait Consultant of: 23andMe, Orion, AsahiKaseiPharma, Pfizer, Collegium Pharmaceuticals.